title: Two conformational states of beta-lactamase bound to GroEL: a biophysical characterization.
abstract: Escherichia coli RTEM beta-lactamase, in which both cysteine residues which form the single disulfide bond have been mutated to alanine residues, can form stable reversible complexes with GroEL under two different sets of conditions. Starting with the GdmCl-denatured enzyme, it is bound to GroEL in a state where no protons are protected against exchange with 2H2O, as determined by electrospray ionization mass spectrometry (ESI-MS). In contrast, when native protein is destabilized at high temperature and added to GroEL, a conformation is bound with 18 protected protons after two hours of exchange. While the high-temperature complex can form both with the wild-type enzyme (with intact disulfide bond) and the Cys-Ala double mutant, only the latter protein can form a complex starting from GdmCl denatured states. Thus, two different sets of conformations of the same protein can be bound, depending both on the conditions used to form the complex and on the intrinsic stability of the intermediate recognized by GroEL, and we have characterized the properties of both complexes.
authors.affiliation: Biochemisches Institut der Universität Zürich, Switzerland.
authors.forename: P
authors.initials: P
authors.lastname: Gervasoni
authors.forename: P
authors.initials: P
authors.lastname: Gehrig
authors.forename: A
authors.initials: A
authors.lastname: Plückthun
mesh_terms: D000409:Alanine / Q000378:metabolism; D018834:Chaperonin 60 / Q000737:chemistry* / Q000378:metabolism; D003545:Cysteine / Q000378:metabolism; D019791:Guanidine; D007700:Kinetics; D046911:Macromolecular Substances; D013058:Mass Spectrometry; D011487:Protein Conformation; D011489:Protein Denaturation; D017510:Protein Folding; D013696:Temperature; D001618:beta-Lactamases / Q000737:chemistry* / Q000378:metabolism*
chemical_list: D018834:Chaperonin 60; D046911:Macromolecular Substances; D001618:beta-Lactamases; D019791:Guanidine; D003545:Cysteine; D000409:Alanine